Ito Chihiro, Haraguchi Ryuma, Ogawa Kohei, Iwata Miku, Kitazawa Riko, Takada Yasutsugu, Kitazawa Sohei
Department of Molecular Pathology, Ehime University Graduate School of Medicine, Shitsukawa 454, Toon, Ehime, 791-0295, Japan.
Department of Hepato-Biliary-Pancreatic and Breast Surgery, Ehime University Graduate School of Medicine, Shitsukawa 454, Toon, Ehime, 791-0295, Japan.
Histochem Cell Biol. 2023 Nov;160(5):407-418. doi: 10.1007/s00418-023-02229-x. Epub 2023 Aug 2.
The liver is known to possess remarkable regenerative potential, but persistent inflammation or severe acute injury can lead to liver fibrosis and incomplete regeneration, ultimately resulting in liver failure. Recent studies have shown that the axis of two types of CXCL12 receptors, CXCR4 and CXCR7, plays a crucial role in liver fibrosis and regeneration. The present study aimed to investigate the regulatory factors involved in CXCR4 expression in injured liver. Immunohistochemical screening of liver tissue samples collected during liver transplantation revealed a reciprocal expression pattern between CXCR4 and MeCP2. An in vitro system involving cultured cell lines and HO treatment was established to study the impact of oxidative stress on signaling pathways and epigenetic alterations that affect CXCR4 mRNA expression. Operating through distinct signaling pathways, HO treatment induced a dose-dependent increase in CXCR4 expression in both hepatocyte- and intrahepatic cholangiocyte-derived cells. Treatment of the cells with trichostatin and azacytidine modulated CXCR4 expression in hepatocytes by modifying the methylation status of CpG dinucleotides located in a pair of TA repeats adjacent to the TATA box of the CXCR4 gene promoter. Only MeCP2 bound to oligonucleotides representing the TATA box region when the cytosine residues within the sequence were methylated, as revealed by electrophoretic mobility shift assay (EMSA). Methylation-specific PCR analysis of microdissected samples revealed a correlation between the loss of CpG methylation and the upregulation of CXCR4 in injured hepatocytes, replicating the findings from the in vitro study. Besides the conventional MEK/ERK and NF-κB signaling pathways that activate CXCR4 in intrahepatic cholangiocytes, the unique epigenetic modifications observed in hepatocytes might also contribute to a shift in the CXCR4-CXCR7 balance towards CXCR4, leading to irreversible liver injury and fibrosis. This study highlights the importance of epigenetic modifications in regulating CXCR4 expression in liver injury and fibrosis.
众所周知，肝脏具有显著的再生潜力，但持续的炎症或严重的急性损伤可导致肝纤维化和不完全再生，最终导致肝衰竭。最近的研究表明，两种类型的CXCL12受体，即CXCR4和CXCR7所构成的轴，在肝纤维化和再生过程中起着关键作用。本研究旨在探讨受损肝脏中CXCR4表达的调控因子。对肝移植过程中收集的肝组织样本进行免疫组织化学筛查，发现CXCR4和MeCP2之间存在相互表达模式。建立了一个涉及培养细胞系和HO处理的体外系统，以研究氧化应激对影响CXCR4 mRNA表达的信号通路和表观遗传改变的影响。通过不同的信号通路，HO处理在肝细胞和肝内胆管细胞来源的细胞中均诱导CXCR4表达呈剂量依赖性增加。用曲古抑菌素和氮杂胞苷处理细胞，通过改变位于CXCR4基因启动子TATA盒相邻的一对TA重复序列中的CpG二核苷酸的甲基化状态，调节肝细胞中CXCR4的表达。电泳迁移率变动分析（EMSA）显示，当序列中的胞嘧啶残基甲基化时，只有MeCP2与代表TATA盒区域的寡核苷酸结合。对显微切割样本进行的甲基化特异性PCR分析显示，受损肝细胞中CpG甲基化的缺失与CXCR4的上调之间存在相关性，这与体外研究结果一致。除了在肝内胆管细胞中激活CXCR4的传统MEK/ERK和NF-κB信号通路外，在肝细胞中观察到的独特表观遗传修饰也可能导致CXCR4-CXCR7平衡向CXCR4方向转变，从而导致不可逆的肝损伤和纤维化。本研究强调了表观遗传修饰在调节肝损伤和纤维化中CXCR4表达的重要性。
